The goal of this work is to study the mechanism of gene regulation in the flagellated kinetoplatida, Leishmania enriettii. The leishmania organism is interesting both as a basic developmental biological system and because of its relevance as an important disease in the developing world. By a basic analysis of gene expression in the leishmania parasite and the identification of developmental regulated genes, we hope to understand both gene regulation in the parasite life cycle and to identify targets for potential disease prevention or treatment. The leishmania parasite is a flagellated protozoa which develops in two distinct environments in nature. In the insect vector, the parasite is extracellular, flagellated and motile and lives in the insect gut. When the insect or promastigote form is introduced into a susceptible host, the promastigote is taken up by host macrophages and undergoes and extensive morphological transformation to the amastigote form. Inside the macrophage, the amastigote is non-flagellated and non-motile. To complete the cycle, when a insect vector bites the infected host, amastigotes are taken up and undergo the reverse transformation to the flagellated, extracellular promastigote form with in the insect gut. Each of these forms of the parasite are functionally and morphologically distinct from one another, each expressing specific genes and modulating the expression of other genes. It is the mechanisms controlling gene expression in these two forms which we intend to investigate. The initial experiments we propose will focus on the regulation of tubulin genes during the amastigote to promastigote transformation. Previous work from other laboratories had demonstrated a marked increase in tubulin protein biosynthesis during the transformation from amastigote to promastigote. We have recently demonstrated that the regulation of the tubulin genes appears to be at the level of RNA transcription and processing. The work proposed is to investigate the molecular basis for this regulation. A second goal of this work is to develop rapid, accurate diagnostic tests for leishmaniasis using specific hybridization of the Leishmania kinetoplast DNA. In previous work we have developed such a method and now propose to refine this technique for the detection of a single Leishmania species using cloned subfragments of the minicircle DNA.